ACTA OTORHINOLARYNGOLOGICA ITALICA 2009;29:119-126

Review

Human Papilloma Virus (HPV) in head and neck

region: review of literature
Linfezione da papilloma virus umano (HPV) nel distretto cervico-facciale:
revisione della letteratura
L. MANNARINI, V. KRATOCHVIL1, L. CALABRESE2, L. GOMES SILVA, P. MORBINI, J. BETKA1, M. BENAZZO
Department of Otolaryngology HN Surgery, University of Pavia, IRCCS Policlinico S. Matteo Foundation, Pavia, Italy;
1
Department of Otolaryngology Head and Neck Surgery, 1st Faculty of Medicine, Charles University, Prague, Czech
Republic; 2 Division of Head and Neck Surgery, European Institute of Oncology, Milan, Italy

SUMMARY
The evidence that human papillomavirus infection is related to head and neck squamous cell carcinoma is supported by molecular and
epidemiological data. The definition of a distinct subset of head and neck squamous cell carcinoma, independent of the traditional risk
factors and with different clinical presentation and outcome, has led to increasing interest in human papillomavirus infection. This review
summarizes current knowledge regarding human papillomavirus biology, oncogenic mechanisms, risk factors for transmission, clinical
significance and prophylactic strategies.

KEY WORDS: Human papillomavirus Head and neck Squamous cell carcinoma Oropharynx Oral cavity

RIASSUNTO
Dati molecolari ed epidemiologici sostengono una correlazione tra infezione da papilloma virus umano (HPV) e carcinomi squamocel-
lulari del distretto cervico facciale (HNSCC). La definizione di un sottogruppo di HNSCC, indipendente dai tradizionali fattori di rischio
e con caratteristiche cliniche e prognostiche differenti, ha portato ad avere un interesse sempre maggiore nellinfezione da HPV. Questa
review riassume le conoscenze attuali sulle caratteristiche biologiche, i meccanismi oncogenetici, i fattori di rischio per la trasmissione ed
il significato clinico e terapeutico dellinfezione da HPV.

PAROLE CHIAVE: Papilloma virus umano Testa e collo Carcinoma squamocellulare Orofaringe Cavit orale

Acta Otorhinolaryngol Ital 2009;29:119-126

Introduction In females, HPV infections, on a global scale, account for

Squamous cell carcinomas (SCCs) represent the most
frequent malignancy in the head and neck region. They
originate from the pluristratified squamous epithelium
which lines the upper aerodigestive tract and are charac-
terised by a multi-phasic and multi-factorial aetiopatho-
genesis 1-10.
Common risk factors in head and neck squamous cell car-
cinoma (HNSCC) are smoking and alcohol abuse, how-
ever, in an increasing proportion of cases, no significant
smoking or drinking history has been reported.
Approximately 35 years ago, a role of human papillo-
mavirus (HPV) in cervical cancer was postulated. Today,
it is well established how this very heterogeneous virus
family represents an important human carcinogen, caus-
ing not only the vast majority of cervical and ano-genital
tumours, but also a variable number of cancers in other
districts of the human body including the head and neck.
more than 50% of infection-linked cancers, in males it is
responsible for barely 5% 11.
HPV positive HNSCC have been reported to share some
epidemiological and biological characteristics 3.
This review will attempt to focus on relevant character-
istics of HPV, analyse its role in oral and oro-pharyngeal
cancer and discuss some emerging developments.
Human Papilloma Virus (HPV) infection
Papilloma viruses are members of the Papillomavirus
family and together with Polyomaviruses form the spe-
cies Papovaviridae. The HPV virion consists of a circular
double DNA strand of about 7.9 kb, protected by a small
capsid. The capsid is about 55 nm in diameter and con-
sists of only two structural proteins. HPV genomes reveal
a well-preserved general organisation. All putative open
reading frames (ORFs) are restricted to one DNA strand.

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L. Mannarini et al.
The second, presumably non-coding strand, contains only
short ORFs which are conserved regardless of localiza-
tion and composition. The individual frames are classified
as early (E) or late (L) genes not unlike other DNA
viruses, where genes are turned on according to a specific
time schedule in the course of a productive infection. The
so-called early genes (E1-E8) are expressed shortly after
infection and prior to the onset of DNA replication. Prod-
ucts of these genes mediate specific functions controlling
replication and expression of viral DNA. In the case of
oncogenic viruses, early gene products are also involved
in the transformation of the host cell. The late genes (L1-
L2) code for the structural proteins of viral capsid and are
activated during the final stages of the viral cycle. Up to 6
early genes and 2 late genes can be detected in HPV 12-20.
HPVs are epitheliotropic viruses that, in the majority of
cases, produce a benign epithelial proliferation. However,
some viral types can be associated with malignant trans-
formation. The genomes of many HPV types have been
re-isolated, sequenced and compared to reference pro-
totypes, countless times, by laboratories throughout the
world. It was found that each HPV type occurs in the form
of variants, identified by about 2% nucleotide differenc-
es in most genes and 5% in less conserved regions. Less
than 100 variants of any HPV type have been detected,
a scenario that is very different from the quasi-species
formed by many RNA viruses. The variants of each HPV
type form phylogenetic trees, and variants from specific
branches are often unique to specific ethnic groups. Im-
migrant populations contain, depending on their respec-
tive ethnic origins, mixtures of variants. Currently, more
than 200 types of HPV are known. The absence of HPV
genomes, intermediate to specific types, show that all
HPV types existed already when humans became a spe-
cies. A growing number of epidemiological, aetiological
and molecular data suggest that variants of the same HPV
type are biologically distinct and may confer differential
pathogenic risks 21.
According to the oncogenic potential, they are classified
as low-risk and high-risk 22. Both high-risk and low-risk
types of HPV can cause the growth of abnormal cells, but
only the high-risk types lead to cancer because only the
E7 protein encoded by high-risk HPVs can immortalize
human epithelial cells. Sexually transmitted, high-risk
HPVs include types 16, 18, 31, 33, 35, 39, 45, 51, 52,
56, 58, 59, 66, 68, and 73 23. It is important to note, how-
ever, that in the genital tract the large majority of high-
risk HPV infections regress on their own and do not cause
cancer 24.
The association between HPV and squamous cell lesions
at various sites of the body, including the oral cavity, was
first described by Syrjnen et al. 25, in 1983. The results
suggested that HPV might be the agent involved in the
development of at least certain special types of oral SCCs
(OSCC) 25. In the next few years, the literature provided
120
further quantitative evidence that oral infection with HPV,
particularly with high-risk genotypes, is a significant in-
dependent risk factor for OSCC 21 26 27.
Oncogenetic mechanism of HPV infection
On the basis of epidemiological and molecular evidence,
in 1995, the International Agency for Research on Cancer
recognized that high-risk HPV types 16 and 18 are carci-
nogenic in humans 28.
It is widely accepted that cancer of the uterine cervix is re-
lated to HPV infection. These two HPV types are respon-
sible for approximately 70% of cervical cancer cases 29.
In addition, high-risk HPVs are associated with other
ano-genital carcinomas, including vulvar, anal, and penile
cancers 30 31 and with some HNSCCs 3.
HPV infection was first held responsible for the develop-
ment of head-and-neck cancer in certain individuals lack-
ing the classical risk factors for this disease (tobacco and
alcohol abuse).
The majority of HPV-related cancers contain HPV DNA
integrated into the host cell genome and express only two
viral genes, E6 and E7, both of which encode oncopro-
teins 32.
Through wounds or abrasions, the papilloma viruses in-
fect the basal epithelial cells that are the only actively di-
viding cells in the epithelium. Virus maturation is closely
related to the degree of epithelial differentiation. Expres-
sion of early viral antigens is found in cells of the basal
layer whereas late viral antigens are formed in the super-
ficial keratinizing epithelial layer 32-36.
The E6 protein of the high-risk HPVs binds and induces
the degradation of the p53 tumour suppressor protein via
an ubiquitin-mediated process, while the high risk E7 may
play a role in the HPV life cycle by disrupting pRB fam-
ily member-mediated transcriptional repression of certain
genes involved in cell cycling. The low-risk HPV-6 E7
protein binds to pRB family members with lower affinity
than that of high-risk HPV-16 and it is not able to immor-
talize the cells 37-41.
Loss of cell cycle and apoptosis control, therefore, con-
stitutes an early and central event in HPV-mediated carci-
nogenesis and the integration of HPV DNA into the host
genome is believed to be the key event 42 43. However, re-
cent studies suggest that transcription of HPV-16 E6/E7
mRNA in tonsillar carcinomas is not necessarily depen-
dent on viral DNA integration and that the viral DNA is
predominately in episomal form 44 and, in this form, also
takes part in the carcinogenesis process 45.
Gene expression profiles of HPV-positive and negative
HN and cervical cancers show substantial differences. A
distinct and larger subset of cell cycle genes is up-regulat-
ed in HPV-positive tumours. Moreover, HPV-positive can-
cers have been shown to over-express testis-specific genes
that are normally expressed only in meiotic cells. HPV-

positive tumours are characterised by the loss of pRb and
Cyclin D1 expression and by over-expression of p16. On
the contrary, HPV-negative over-express pRB and Cyclin
D1, and under-express p16. These findings emphasize the
potential value of targeting E6 and E7 protein. The prod-
uct of the E6 gene binds wild-type p53 and induces its
degradation. The loss of functional p53 impairs apoptosis
and induces genetic instability. Moreover, E6 inactivates
telomerase, an enzyme that maintains telomeric DNA sta-
bility. Protein E7 binds retinoblastoma protein (pRb) and
other related proteins. In this way, it causes the release of
transcriptional factors that activate genes regulating cell
proliferation. pRb down-regulation by HPV E7 results in
p16 up-regulation46-54.
At least in SCCHN, HPV-induced genomic instability
does not necessarily lead to cancer. There are many factors
involved in carcinogenesis, HPV infection being only one
of them. However, HPV high risk serotypes are detected
3-fold more frequently in patients with malignant lesions
than in those with benign or pre-cancerous lesions54.
Risk factors for HPV infection and cancer
Epidemiological studies on cervical cancer have clearly
demonstrated that high-risk mucosatropic HPVs are trans-
mitted by sexual contact 28. The means by which HPV is
transmitted to the upper airways is unclear. Oral HPV in-
fection is rare in newborn babies of infected mothers 55
and in children prior to sexual activity 56; infections in-
crease after onset of sexual activity 57.
It is generally assumed that HPV infection precedes the
development of HPV-positive HNSCC: the presence of
high risk HPV infection in the oral mucosa and seroposi-
tivity significantly increase the risk of development of
OSCC 58-61. Therefore, risk factors for HPV oral infection
are likely, by extension, to be risk factors for HPV posi-
tive HNSCC.
Patients with HPV-positive tumours appear to be distinct
from HPV-negative patients. While tobacco-associated
OSCCs are more frequent in men, men and women are at
equal risk of HPV-associated OSCC. In addition, patients
with HPV-associated OSCC are often non-smokers and
non-drinkers 62-64 and younger than patients with HPV-
negative tumours 65.
Although evidence suggests that HPV is associated with
cancer in non-smokers and non-drinkers, the degree to
which oral HPV infection may combine with tobacco
and/or alcohol use to increase the risk of cancer is un-
clear. Controversial data exist, suggesting a concurrent
action, either as a synergistic (multiplicative) 66 or as an
additional effect 2.
Several case-control studies have reported that certain
types of sexual behaviour increase the risk of OSCC.
Risk factors, among men, include young age at first inter-
course, number of sexual partners and history of genital
Human papilloma virus (HPV) in head and neck region
warts. Risk is increased in women with a large number of
sexual partners 66-69.
Furthermore, some investigations have demonstrated that
certain types of sexual behaviour are strongly associated
with the risk of HPV-positive tumour, including history
of oral sex and oral-anal contact 70 71. An increased risk
of HPV-associated OSCC in individuals with a history
of HPV-associated ano-genital cancers and in husbands
of women with in situ carcinoma and invasive cervical
cancer confirms oral-genital transmission, although direct
mouth-to-mouth contact could not be excluded as a means
of transmission 72.
The role of marijuana use in HPV-associated OSCC is
still unclear, as well as the relationship with poor oral hy-
giene; a recent study reported that HPV-positive cancers
were independently associated with sexual behaviour and
exposure to marijuana but not related to tobacco smoking,
alcohol drinking, and poor oral hygiene 73.
Increased risk for OSCC seems to be related also to im-
muno-suppression. Recent evidence has indicated that
HPV-related diseases are increased in the oral cavity of
human immunodeficiency virus (HIV)-positive individu-
als 74 and new data imply that the problem of HPV-related
cancers do not decrease in HIV-positive men and women
in the anti-retroviral therapy era 75; genetic susceptibility
such as that in patients with Fanconi anaemia also increase
the risk of HPV-mediated tumourigenesis 61.
Oral and oro-pharyngeal squamous cell
carcinomas related to HPV infection and
clinical meaning
Oral and oro-pharyngeal squamous cell carcinoma is a
disease usually related to environmental exposures, pri-
marily tobacco and alcohol abuse. However, 15-20% of
these cancers occur in patients without traditional risk
factors 3 61-66 76.
Data appearing in the literature have provided strong evi-
dence that HPVs may be the cause of a defined subset of
head and neck cancers, particularly those of the oro-phar-
ynx (tonsils and throat) and also an indicator of improved
survival. The International Agency for Research on Can-
cer conducted a multicentre case-control study on oral
cavity and oro-pharynx carcinomas, in 9 countries 61. Of
these tumours, 70% harboured HPV DNA. HPV16, most
commonly observed in genital cancers, was also the most
common type found in these tumours. The study conclud-
ed that HPV appears to play an aetiologic role in many
cancers of the oro-pharynx and possibly a small subgroup
of cancers of the oral cavity 61.
Some studies aimed at defining the molecular profile of
HPV-positive OSCC. In a recent study, OSCC HPV-posi-
tive cancers with a high p16 expression showed signifi-
cantly better response, independently of the treatment
(surgery vs organ-sparing) 62. However, at present, data on
121
L. Mannarini et al.
immunohistochemical markers of radiochemiosensitivity
are controversial. An organ-sparing trial on advanced oro-
pharyngeal cancer demonstrated that low EGFR and high
p16 (or higher HPV titre) expression were favourable
predictive markers in outcome 77 whereas a retrospective
study concerning surgically treated OSCC, characterized
by the same immuno-histochemical profile (HPV posi-
tive/high p16 expression) showed that p16 over-expres-
sion is not a prognostic marker of relapse and second tu-
mours 78.
The presence of antibodies against HPV, in the serum,
should be an easily accessible marker of virus detection 79.
The human body produces antibodies against viral cap-
side proteins, against early viral proteins, including onco-
proteins E6 and E7. The relationship between the levels
of antibodies and presence of HPV positive tumour cells
could be used in early diagnosis 60, monitoring the course
of the disease and early detection of recurrence of oral
and oro-pharyngeal cancer 59 78 80. Approximately 10% of
healthy individuals develop a persistent infection and it
is this cohort that should be monitored as being at risk of
cancer progression 81.
In the majority of the studies, OSCC related to HPV in-
fection shows a better outcome and a reduced risk of re-
lapse and second tumours compared with HPV-negative
tumours 2 59-61 82 83. The concept of better treatment results
in HPV-positive tumours has been confirmed both in pa-
tients submitted to organ-preservation treatment as well
as in those submitted to surgery 82-84. The reason for the
improved survival is unclear. However, better outcomes
seem to be attributed to the ability of HPV-positive cancer
cells to induce apoptotic cell death in response to DNA
damage 85. Another reason appears to be attributed to the
absence of carcinogen-induced early genetic changes in
the epithelium and the development of multifocal tumours
(field cancerization concept) 86.
The enhanced radiochemiosensitivity reported in the lit-
erature highlights the need of meticulous clinical trials
to determine optimum management of HPV-associated
OSCC, compared with HPV-independent OSCC 87 88.
A better understanding of HPV-associated carcinogen-
esis is necessary for the development of HPV-targeted
strategies. In the absence of cell mutations, classically
associated with carcinogenesis in tobacco-associated
oropharyngeal cancers, the repression of E6 and E7 ex-
pression, in HPV-positive carcinomas, results in restora-
tion of Rb and p53 tumour-suppressor pathways and is
sufficient to arrest cell growth or cause apoptosis 89 90. In
fact, HPV-associated cancers constantly express the HPV
E6 and E7 viral oncogenes, even in the late stages of the
disease, and repression of viral oncogene expression by
drugs that interfere with the expression or function of vi-
ral proteins can induce senescence or apoptosis of cancer
cells. Moreover, therapeutic vaccines that elicit a cytolytic
immune response to cells expressing viral proteins could
122
be curative, possibly even in the advanced stages of the
disease 89 90.
As far as diagnostic implications are concerned, it has been
postulated that the identification of HPV in metastatic cer-
vical lymph nodes could be used to assess the tonsillar
origin of metastatic carcinomas with an unknown primary
tumour. HPV-positive metastases have been identified not
only by ISH for HR-HPV and p16 immuno-reactivity 91 92
but also by the strong correlation with a distinct non-kera-
tinizing morphology 93 94.
Vaccination
Vaccines designed strictly for prevention of cervical can-
cer and vulvar genital warts have been introduced over the
last few years. To date, there are two commercial vaccines,
quadrivalent Gardasil (registered trademark of the Merck
& Co., Inc., USA) protects against HPV types 6, 11, 16,
18 and bivalent vaccine, Cervarix (registered trademark
of the GlaxoSmithKline Group of Companies, Australia)
which targets HPV types 16 and 18. Experimental studies
with L1-VLP vaccines showed their ability to induce nat-
ural immunity. Antibodies are a primary defence against
HPV infection. Both existing vaccines are able to create a
robust humoral immune response 95 96 which is much more
effective than the levels of antibodies that can be acquired
after a natural infection, and persist for a period of at least
60 months 97. A 5-year follow-up demonstrated 100%
effectivity in the prevention of persisting infection and
HPV-16 and HPV-18 CIN 2/3 lesions in young women.
Oral HPV infection seems to be the main risk factor for
cancerogenesis of HPV-positive oro-pharyngeal cancer
and HPV 16 (type contained in both above-mentioned vac-
cines) is found in the majority of cases of HPV positive oral
cancer 98. For this reason, it might be possible to prevent or
even treat these cancers by means of vaccines designed
to elicit appropriate virus-specific immune responses. It
is tempting to suggest that if high-risk HPV infection is
prevented, the subsequent development of invasive cancer
caused by HPV will be abolished. The impact of current
HPV vaccines on the incidence of persistent oral HPV in-
fection remains to be identified. Clinical trials to evaluate
the efficacy of the quadrivalent HPV vaccine in protecting
against oral HPV infection are currently underway.
HPV infection is, generally, sexually acquired 99, and,
therefore, vaccination should be performed prior to sex-
ual debut in order to prevent serious HPV-related genital
and oral diseases. All vaccine trials reported to date have
been designed to investigate the ability of the vaccines to
generate protection against the consequences of ano-geni-
tal HPV infection in women. However, there is reason to
be optimistic that the existing vaccines may be protective
against oral HPV infection, and, therefore, effective in
preventing vaccine-type HPV-associated head and neck
cancer both in men and women. Although vaccination

currently involves exclusively the female population, im-
munogenicity studies have demonstrated that the vaccines
elicit a robust humoral immune response also in men, an
important finding considering that the majority of HPV-
associated head and neck cancers occur in men. Thus, it is
possible that an HPV vaccine could have benefits beyond
the target population 94-104.
Data on oral HPV infections, in the immunization setting,
are limited to animal models, which have shown a protec-
tive effect and a reduction in the development of HPV-re-
lated oral lesions 105 106. These data imply that therapeutic
vaccines will likely be effective for low-volume disease.
Therefore, these vaccines could be used as adjuvant treat-
ment following surgery or radiotherapy to clear microscop-
ic residual disease by generating an immune response.
Conclusion
In recent years, there has been an increase in the annual
incidence of HPV-related HNSCC in the United States
and Europe 107.
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